Method for the extraction of lysergol and ergot alkaloids from plants of the ipomoea genus

ABSTRACT

A method is disclosed for recovering ergolic alkaloids from the seeds of the Kaladana plant, the latter being a plant of the Ipomoea section of Convolvulaceae, Calonyction genus (Choisy) Hallier f, species nova. The method is a selective extraction with appropriate solvents, in order to recover both lysergol and chamoclavine from the Kaladana seeds. Temperatures and times of treatment are critical and the preferred solvents are chlorinated aliphatic hydrocarbons.

United States Patent [1 1 [Ill 3,920,663 Ferrari [45] Nov. 18, 1975 [54'] METHOD FOR THE EXTRACTION 0F 3.224.945 l2/l965 r |r.................. 260/285.5

LYSERGOL AND ERGOT ALKALOIDS FROM PLANTS OF THE IPOMOEA GENUS Inventor: Giorgio Ferrari. Milan. Italy Assignee: Simes S.p.A.. Milan. Italy Filed: Aug. 14, 1972 Appl. No.: 280.2l2

Foreign Application Priority Data Aug, l7. l'47l Belgium H 'IIJR Jan. l-l. W71 Belgium r. IlISSb US. Cl 260/2855: 260/3l3.l Int. Cl. C07D 519/02 Field of Search 260/2855, 3l3.l

References Cited UNITED STATES PATENTS 4/]938 Kuessrnen 260/285.$ 9/l94l Moore Zoo/285.5

l0ll957 260/2855 M11965 Rutschmann et al ISO/285.5

OTHER PUBLICATIONS Merck Index. 8th Edition. p. 228 (1968).

Erge et al.. Chem. Abstr. Vol. 79. Col. 396006 (I973).

Voigt et al., Die Phurmuzie. Vol. 26. pp. 494403 503 l97l Primary Examiner-Donald G. Daus Armmqv, Agent. or Firm-Shlesinger. Fitzsimmons 8.: Shlesil'lger l 5 7 I All STRA (T A method is disclosed for recovering crgolie alkaloids from the seeds of the Kaladana plant. the latter being a plant of the lpomoca section of Convolvulaceac. Calonyction genus (Choisy) Hallier f., species nova. The method is a selective extraction with appropriate solvents, in order to recover both lysergol and chamoelavine from the Kaladana seeds. Temperatures and times of treatment are critical and the preferred solvents are chlorinated aliphatic hydrocarbons.

13 Claims, 3 Drawing Figures US. Patent Nov. 18. 1975 METHOD FOR 'l'llli I-ZX'I'RAG'I'IUN (ll [.YSERGUI. AND l-ZRtlUT ALKALOlDS FROM PLANTS OF THE llOMOlI/l (ilINllS 'l'his invention relates to the extraction of alkaloids having an ergolic structure from a plant of the lpouroea genus. (Convolvulaceae family).

More detailedly'. the present invention relates to the extraction and isolation in the pure condition of lysergot and chanoclavine from the seeds of Kaladana. a plant identified by tlte scientific name Calonyction lpomoea (Cltoisy) llallier f.. nova species.

Kaladana is the native name by which is indicated a spontaneous plant of the subtropical mountainous territories. more particularly the pre-Hintalayan belt tlndial. whose scientific classification has been controversial and noticoncordaut heretofore;

As a matter of fact iii the publication Chopra's Indigenous Drugs ofludia. UN. Dhur and Sons l'rivate l itlt' ited. Calcutta. I958. page Sl. Kaladana is indicated as corresponding to the scientific term lprmtoeu Iredvracen (LP).

ContraryWise. in J. Pharm. Sc.. Vol. 56. page HI. 1967. in the context of an article by Gupta and coworkers. it is afl'rrmed that Kaladana is the name by which Iprmwea rrrrrrr'cara is commonly known. Lastly. in the "ltritish Pharmaceutical Code i949." page 459. it is affirmed. conversely. that Kaladana is composed by dried seeds of lpomoea lrederaceal [border to dissipate any possible doubt. it is deemed fitting to describe it more comprehensively.

Kaladana as of interest in the present invention is a plant which certainly-belongs to the Convolvulaceae family. section lpornoeae and genus Calonyetion. which exhibits the following features:

- A lianous perennial plant having voluble twines which are very long. branch-like. herbaceous but wood-like hardened at the base. furrowed by two longitudinal grooves which are abundantly fitted with short herbaceous thorns. these beir'ig slightly curved down wards and having an obtuse apex. non-stinging.

Akin to (.murr'camm (L) G. Don. it differs therefrom forthe leaves which are alternate and ehordate-acuminate. neither lobate nor sagittate. having slightly sinuateundulate edges and a greater size: length up to about 25 ems. and width up to 22 cms.. the petiole being long from A to '16 the length of the leaf. shortly expanded at the base and scarcely hairy there. I

The flowers are in number of 2 3 on an axillar peduncle; the coral is elongate conical tubular. whitish. having a length shorter than that of (.murt'catrrm (about 7 cms.t. funnel-like rotated edge. which is obutusely pentalobate tntd has a diameter of about it ems. the colour being lilac or rosy. tttore or less dark. which opens at night time. stamina and stylus do not protrude from the eorol tube. the calix has aetuuinate and a rplicate lobes. which however become enlarged until becoming ovate-acuminate and slightly divergent as the capsule ripcns.

The capsule is pendulous. ovate-articulate having three tfrom two to four) black seeds in the form of a clove (outer face nvoidal and the internal faces plaoar). minutely tomentous-granulous. long 7 ntms. as an a erage and wide 6 mms.

As' to its habitat. the plant is spontaneous in the nu'nrntainous rue-Himalayan belt of India and Pakistan (l-Iast l. up to an altitude of I50" meters. The drug. conill (ill

2 -isting of the seeds. is locally used for its aperrent action. whereas the powder is used as all arttipyretic.

In the. accompanying drawings. which show Kaladana. there are shown:

In l-'l(i. l a twine of the plant under discussion with leaves and flowers;

In FIG. 2 the seed-carrying capsulae when ripened.

and

In FIGS. 3 and 4 the front and side views of a seed.

respectively.

lt has now been found that this plant. and more particularly its seeds. are particularly rich with ergolic alkaloids and these can be usefully extracted by a method which is comparatively simple and allows. inter alia. to obtain them in a pure condition.

According to the invention. the Kaladana seeds are ground until obtaining a flour The ground drug is subseqnctttly cshaostcd in the cold with gasoline or light naphtha in order to return e all the fat portion The op elation is repeated until complete exhaustion is ohtained. that which is achieved by repeating the operation three or four times.

The fat-stripped drug is then subjected to an extraction with appropriate solvents for withdrawing the active prittciples. Especially useful to this end are the halogen substituted aliphatic hydrocarbons such as chloroform. carbon tetrachloride. methylene chloride and so forth. In order to facilitate the extraction. the halogen substituted aliphatic hydrocarbon can be supplemented by a small percentage of a low molecular weight aliphatic alcohol and a small amount of an bydroxide. Alcohols such as methanol. ethanol. isopropanol and hydroxides such as ammonia are very well suited to this purpose. The amounts of alcohol added can attain ISZ whereas the alkali metal bases cart reach 5% by volume with respect to the chlorinated hydrocarbon. The extractions of the material are carried out. preferably. at a temperature comprised between' lO'C and 50C. The number of extractions which are required for a complete exhaustion of the alkaloids of the drug is from three to Five.

The extracts as obtained are subsequently evaporated to a volume which is approximately equal to one tenth of the original one at a temperature'of 30C and under su'batmospherical pressures and allowed to stand a few days at OC- lC. The mass which separated after this stay is removed by filtration. The precipitate cake. in turn. is slurricd in ten times its weight of water and the portion which is insoluble is dissolved in methanol and combined with the filtrate. The filtrate is washed with water. v i

From the organic phase. which is washed with water. the extraction of ergolic alkaloids is proceeded with by osing aqoeous solutions of acids. lt.has been found that the aqueous diluted solutions of phosphoric acid lend themselves very well to the task. The extraction of the organic phase with aqueous solutions of acids is continued until the liltrlielt alkaloid testbecomes negative.

l-rom thecombined acidic extracts. which are made weakly alkaline with a base (preferably aqueous ammouial. the alkaloids are extracted again with a mixture of an aliphatic chlorinated hydrocarbon and an aliphatic alcohol. A mixture of chloroform and methanol in thc ratio of 7:3 by volume proved to be particularly satisfactory.

The extractions are repeated until the Ehrlich test is negative. The combined extracts are washed once with water. dried over anhydrous sodium sulphate and then 3 evaporated to dryness at a temperature below 30C under subatmospherical pressures.- The residue is formed by tlte alkaloid fraction'ofKaladana. that is lpomoea Calonyction (Choisyl Hallie r f. nova species and.

when examined in thin-layer chromatography. by using as the solvent substance ntethylene chloride-mcthanoh benzene-t25z$z5i and as the detector at 3% solution in alcohol of vanillin and 0.5% by volume of concentrated sulphuric acid.aftcractivation at llOC IC during 5 minutes. exhibits the presence of several crgolie alkaloids. among which lysergol predontinates. and. secondly. chanoclavine.

lt has been found tltat lysergol can be isolated by the total alkaloids by merely washing the mass with a low molecular weight aliphatic alcohol such as methanol. andanakes up the residue which is insoluble in the cold. Such a residue. upon filtration. can be further purified by crystallization front an appropriate organic solvent. or also from a mixturc of dimcthylsulphosidc attd water. Lyscrgol is especially well purified by repeated crystallizations from dimethylsulphoxide water in the ratio of lzl: the product thus obtained exhibits the following properties: the percentage analysis corresponds to the raw formula C,,H,,.ON,.

C'7r Calcd. 75.55; H'7r Calcd. 7.14: N7l Caled. 11.02.

Found: 75.39: Found: 7.22: Found: 1.96.

mol.wt. 254.3; m.p. (crystallised from alcoholl253- if-255C. (decomp.). lai +54. (t -=03 'in pyridine). i

According to another aspect. the present invention provides for the further isolation ofchanoclavinc front the residue which contains the total ergolic alkaloids. upon isolation of lysergol. To thisend the present invention provides for the further steps of:

a. evaporation to dryness. in a vacuo and at a temperature below 60C. of the mother liquors which are left after the extraction of lysergol;

h. dissolution of the residue iii an excess of pyridine and addition of acetic acid anhydride in an amount by weight equal to that of the dried residue. allowing the mixture to stand during 24 hours;

c. precipitation of the chanoclavinc diaeetate by pouring the reaction product obtained in an excess of water and ice. and

d. purification of the chanoclavine diacetatetrawl after drying by recrystallization from ethyl acetate.

The thusly obtained chanoclavine diacetate has the following properties: i

melting point l72C-l 73C [0],, --53.4(=(l.95 in pyridine): [01],, --$8 (c=lZ in chloroform).

LR; spectrum in CHCl characteristic absorption bands at I625 cm (N-COCH;r); i730 cm" (O-COCH UN. spectrumin McOll lX CHCl A max 283 not (log 1 ,Htl 2th out (loge 4.76l;.2)l not (log i Analysis for C:,,l-l, .O N, (340.4)

Found: 70.3; M5; 8.3. i i

As regards the stages of the ntcthod outlined above it should be recalled that:

l. The dry residue as obtained upon evaporation of the tnotlter liquors combined of the Iysergol extraction. is weighed and dissolved. preferably in about three times' its weight of anhydrous pyridine;

2.,The excess ofwater and ice for the precipitation of chanoclavine diaeetate is about ten times the volume of the reaction product of the stagc(b);

3. The precipitate of chanoclavine diaectate (raw) is separated by filtration upon repeated washings with water and then purified.

The lysergol obtained according to the present invention. it has been found. exhibits interesting phsiological properties which manifest themselves on the test animals on different systems and especially on the nervous system and the circulatory system. The substance. in very low dosages. acts as a hypotensivc which is particularly active as to 'its effect on the peripheral circulatory system. The substance exhibits considerable psychotropic properties with antiserotonin effects. The substance exhibits pronounced analgesic properties. Lastly. the stimulating effect on isolated organs. such as uterus and intestine. is considerable. Lysergol. on account of lltese features cart find a useful application in the therapeutics for different pathological situations. To this end it could be formulated in an appropriate manner. as such. or in the form of soluble salts with appropriate excipients suitable for both parcnthcral and oral use.

Examples are now given by way of example only and without any lintitation.

EXAMPLE I kgs. of Kaladana seeds are ground in a ntill so as to obtain a flour having a size of from 40 to meshes per cm.

The ground drug is exhausted in the cold condition in an extractor equipped with a stirrer. live times with gasoline having a boiling point oftitlC C. 150 liters of solvent are employed each time. Every extraction step takes three hours with stirring. The det'atted drug. upon complete removal of the gasoline. is extracted in the same apparatus twice with a mixture of chloroformmetbanoi-antmon'ia in the ratios 9:013:01. I60 liters of the mixture are employed.

Subsequently. the mass is extracted three additional litnes with chloroform only liters).

.The combined extracts are evaporated under subatmospherical pressures at a temperature below 30C until obtaining a volume of 25 liters.

The concentrate is allowed to stand in a refrigerator at 0C 4'(. during two days. The solid which has been separated is collected by filtration with a suction putnp and the cake is dried under vacuum. The dried cake (300 to 400gramsl is slurried cold in If) times its weight of water and stirred during one hour. Filtration is repeated and the water insoluble residue is dissolved in thrice its weight of methanol; the methanol solution being added to the main filtrate. The filtrate is now washed with (r liters of water in a separator. The organic pltasc is theii further evaporated until a volume of eight liters is attained. in a vacuo and at a temperature below 30%. This concentrate is exhausted by extracting it four times with a 5% solution of phosphoric acid. There is employed. in total. eight liters of the acidic solution. The combined acidic extracts are made slightly alkaline with aqueous ammonia until reaching a pH of Rand are exhausted by extracting them with a chloroform-methanol mixture in the ratio 7:3. Four extractions are carried out by employing. as a total. 8 liters of the mixture.

' The organic extracts are washed once with water. then they are dried over anhydrous sodium sulphate andevaporated to dryness. under subatmospherical pressures at a temperature below 30C The residue (200 to 27!),grams) as formed by the total crgolic alkaloids oi the drug has the composition indi 'ated in the introductory part hereof and is now stirred in the cold with 500 grants of methanol during 1 hour. The insoluble portion is collected on a filter and dried in a vacuo.

The further purification of the insoluble fraction (lysergol) is carried out by dissolving the raw product in an equal amount by'weight oi dimethylsulphoxide. by heating gently on a water bath. The solution is treated with decolorizing charcoal. filtered carefully and the filtrate is supplemented with an equal volume of distilled water. Then it is allowed to stand for crystallizing. The cr'ystallizate is separated by filtration and dried in a vacuo to constant weight.

The operation is repeated until obtaining a product having the properties as indicated in the introduction hereof.

sxmuru; z

The metltanolic mother liquors as obtained from the washing of the ergolic total alkaloids of Example I are placed ina rotary evaporator and the solvent is continuously removed in a vacuo to dryness at a temperature below 60C. The residue (70-90 grams) is taken up with anhydrous pyridine (2l0 270 mls.) and the solution is supplemented with acetic acid anhydride (70-90 mls.).

The mass. sheltered from moist air. is allowed to stand during 24 hours. Subsequently is poured over water and ice (2000 2200 mls.). The product which separates is allowed to stand and is repeatedly washed with water. I t

it is then ,i'iltercd under vacuntm. dried and recrystallized l'rom'ethyl acetate.(3O 4S grs.). The thusly obtained chanoclavine diacetate is pure when examined 7 in thin layer chromatography on a Kieselgcl GP, layer (Type 60) by using methylene chloride-benzenemethanol 25/5/5 as the solvent and vanillin and sulphuric acid as the reagent.

What is claimed is: g

l. A method for the preparation oi lysergol and ergolic alkaloids comprising the steps of extracting these alkaloids with a solvent from the seeds of the plant Kaladana. that is. a plant of the family Convolvulaeeae. section of lpomoeac. genus Calonyction (Choisy) Haliier i. nova species. and concentrating the extract.

2. A method according to claim I. wherein the seeds oi the plant are ground to a flour and fat-stripped and then extracted with a halogen-substituted aliphatic hy- .drocarbonaceous solvent selected from the group con-- sisting of chloroform. carbon tetrachloride. and methylene chloride. and at a temperature front iOC to 50C. the extraction step being repeated from 3 to 5 times.

J. A. method according to claim 2. wherein the solvent is supplemented by an amount. not exceeding lS'Z by volume with respect to the solvent. ola low molecular weight aliphatic alcohol selected from the group consisting of methanol. ethanol and isopropanol. anti an alkali metal in an amount. not exceeding 5% by volume with respect to the solvent.

4. A method according to claim 2. wherein the solvent is supplemented by a liquid selected from the group consisting oi methanol. ethanol anti isopropanol and ammonium hydroxide.

5. A method according to claim 3. whcrein the ex tracts are combined and concentrated to one tenth of their original volume at a temperature below 30C under sub-atmospheric pressure. are allowed to stand a few days at 0C 4C and subsequently filtered. the liltrate is washed with water. the filtrate is made alkaline. and is extracted with an aqueous acidic solution. to separate the ergotic alkaloids from the contbined acidic extracts. said latter extraction being continued until the Ehrlich alkaloid test is negative. the alkaloids are extracted again with a mixture 0| a chlorinated aliphatic hydrocarbon and an aliphatic alcohol. the new extraction being continued until said Ehrlich test is negati e again. and the combined extracts. washed once only with water and dried over anhydrous sodium sulphate are evaporated to dryness at a temperature below 30C under sttbatntosphcric pressure. forming a residue composed of the alkaloid fraction oi the extracted plant. itt which iyscrgol predominates.

6. A method according to claim 5. wherein said aqueous acidic solution is a solution of phosphoric acid.

A method according to claim 5. wherein said mixture [oi the second extraction stage is a mixture of chloroform and methanol in the respective volume ratio of 7:3.

8. A method according to claim 5. wherein tlte acidic extracts are made alkaline with aqueous ammonia before the second extraction stage.

9. A method according to claim 5. wherein lysergol is isolated from said residue by washing with a low molecular weight aliphatic alcohol. such as methanol. filtered. and further purified by crystallization with a solvent selected from the group consisting olorganic solvents and a mixture ol'dimcthyisulphoxide and water in a l:l raiio.

10. A ptethod for the extraction of chanoclavine. wherein the mother liquors. which are left from the lysergol isolation step as claimed in claim 7. are evaporated to dryness under vacuum and at a temperature below 60C. the dry residue being dissolved in an excess of anhydrous pyridine. and acetic acid anhydride is added thereto in an amount by weight equal to that oi the rcsidue.-the reaction mixture being allowed to stand during) hours. the chanoclavine diaeetate being precipitated by pouring the mixture as obtained from the reactidnin an excess of water and ice. the raw chanoclavine diacetate being purified after drying it. by recrystallisation from ethyl acetate.

.1 I. A method according to claim it]. wherein the excess of anhydrous pyridine is about three times the weight ol' the dry residue.

l2. A method according to claim I0. wherein the excess of water and ice is about ten times the volume of tlte reaplion product.

1.1. A method according to claim I. wherein a fraction of the plant is dissolved in a halogen substituted aliphatic hydrocarbon solvent. the extract is evaporated and liltered. the filtrate is then extracted with an aqueous acidic solution. the acidic extract is then made slightly alkaline. and the alkaloids so formed are again extracted with a mixture oi an aliphatic chlorinated hydrocarbon ttnd an aliphatic alcohol. and then washed and dried.

Q O i i 

1. A METHOD FOR THE PREPARATION OF LYSERGOL AND ERGOLIC ALKALOIDS COMPRISING THE STEPS OF EXTRACTING THESE ALKALOIDS WITH A SOLVENT FROM THE SEEDS OF THE PLANT KALADANA, THAT IS, A PLANT OF THE FAMILY CONVOLVULACEAE, SECTION OF IPOMOEAE, GENUS CALONYCTION (CHOISY) HALLIER F. NOVA SPECIES, AND CONCENTRATING THE EXTRACT.
 2. A method according to claim 1, wherein the seeds of the plant are ground to a flour and fat-stripped and then extracted with a halogen-substituted aliphatic hydrocarbonaceous solvent selected from the group consisting of chloroform, carbon tetrachloride, and methylene chloride, and at a temperature from 10*C to 50*C, the extraction step being repeated from 3 to 5 times.
 3. A method according to claim 2, wherein the solvent is supplemented by an amount, not exceeding 15% by volume with respect to the solvent, of a low molecular weight aliphatic alcohol selected from the group consisting of methanol, ethanol and isopropanol, and an alkali metal in an amount, not exceeding 5% by volume with respect to the solvent.
 4. A method according to claim 2, wherein the solvent is supplemented by a liquid selected from the group consisting of methanol, ethanol and isopropanol and ammonium hydroxide.
 5. A method according to claim 3, wherein the extracts are combined and concentrated to one tenth of their original volume at a temperature below 30*C under sub-atmospheric pressure, are allowed to stand a few days at 0*C - 4*C and subsequently filtered, the filtrate is washed with water, the filtrate is made alkaline, and is extracted with an aqueous acidic solution, to separate the ergotic alkaloids from the combined acidic extracts, said latter extraction being continued until the Ehrlich alkaloid test is negative, the alkaloids are extracted again with a mixture of a chlorinated aliphatic hydrocarbon and an aliphatic alcohol, the new extraction being continued until said Ehrlich test is negative again, and the combined extracts, washed once only with water and dried over anhydrous sodium sulphate are evaporated to dryness at a temperature below 30*C under subatmospheric pressure, forming a residue composed of the alkaloid fraction of the extracted plant, in which lysergol predominates.
 6. A method according to claim 5, wherein said aqueous acidic solution is a solution of phosphoric acid.
 7. A method according to claim 5, wherein said mixture for the second extraction stage is a mixture of chloroform and methanol in the respective volume ratio of 7:3.
 8. A method according to claim 5, wherein the acidic extracts are made alkaline with aqueous ammonia before the second extraction stage.
 9. A method according to claim 5, wherein lysergol is isolated from said residue by washing with a low molecular weight aliphatic alcohol, such as methanol, filtered, and further purified by crystallization with a solvent selected from the group consisting of organic solvents and a mixture of dimethylsulphoxide and water in a 1:1 ratio.
 10. A method for the extraction of chanoclavine, wherein the mother liquors, which are left from the lysergol isolation step as claimed in claim 7, are evaporated to dryness under vacuum and at a temperature below 60*C, the dry residue being dissolved in an excess of anhydrous pyridine, and acetic acid anhydride is added thereto in an amount by weight equal to that of the residue, the reaction mixture being allowed to stand during 24 hours, the chanoclavine diacetate being precipitated by pouring the mixture as obtained from the reaction in an excess of water and ice, the raw chanoclavine diacetate being purified after drying it, by recrystallization from ethyl acetate.
 11. A method according to claim 10, wherein the excess of anhydrous pyridine is about three times the weight of the dry residue.
 12. A method according to claim 10, wherein the excess of water and ice is about ten times the volume of the reaction product.
 13. A method according to claim 1, wherein a fraction of the plant is dissolved in a halogen substituted aliphatic hydrocarbon solvent, the extract is evaporated and filtered, the filtrate is then extracted with an aqueous acidic solution, the acidic extract is then made slightly alkaline, and the alkaloids so formed are again extracted with a mixture of an aliphatic chlorinated hydrocarbon and an aliphatic alcohol, and then washed and dried. 